Department+of++Chemical+and+Biomolecular+Engineering
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3D Visualization and Quantification of Bioplastic PHA in a Living Bacterial Cell
3D holographic microscopy leads to in-depth analysis of bacterial cells accumulating the bacterial bioplastic, polyhydroxyalkanoate (PHA)
A research team at KAIST has observed how bioplastic granule is being accumulated in living bacteria cells through 3D holographic microscopy. Their 3D imaging and quantitative analysis of the bioplastic ‘polyhydroxyalkanoate’ (PHA) via optical diffraction tomography provides insights into biosynthesizing sustainable substitutes for petroleum-based plastics.
The bio-degradable polyester polyhydroxyalkanoate (PHA) is being touted as an eco-friendly bioplastic to replace existing synthetic plastics. While carrying similar properties to general-purpose plastics such as polyethylene and polypropylene, PHA can be used in various industrial applications such as container packaging and disposable products.
PHA is synthesized by numerous bacteria as an energy and carbon storage material under unbalanced growth conditions in the presence of excess carbon sources. PHA exists in the form of insoluble granules in the cytoplasm. Previous studies on investigating in vivo PHA granules have been performed by using fluorescence microscopy, transmission electron microscopy (TEM), and electron cryotomography.
These techniques have generally relied on the statistical analysis of multiple 2D snapshots of fixed cells or the short-time monitoring of the cells. For the TEM analysis, cells need to be fixed and sectioned, and thus the investigation of living cells was not possible. Fluorescence-based techniques require fluorescence labeling or dye staining. Thus, indirect imaging with the use of reporter proteins cannot show the native state of PHAs or cells, and invasive exogenous dyes can affect the physiology and viability of the cells. Therefore, it was difficult to fully understand the formation of PHA granules in cells due to the technical limitations, and thus several mechanism models based on the observations have been only proposed.
The team of metabolic engineering researchers led by Distinguished Professor Sang Yup Lee and Physics Professor YongKeun Park, who established the startup Tomocube with his 3D holographic microscopy, reported the results of 3D quantitative label-free analysis of PHA granules in individual live bacterial cells by measuring the refractive index distributions using optical diffraction tomography. The formation and growth of PHA granules in the cells of Cupriavidus necator, the most-studied native PHA (specifically, poly(3-hydroxybutyrate), also known as PHB) producer, and recombinant Escherichia coli harboring C. necator PHB biosynthesis pathway were comparatively examined.
From the reconstructed 3D refractive index distribution of the cells, the team succeeded in the 3D visualization and quantitative analysis of cells and intracellular PHA granules at a single-cell level. In particular, the team newly presented the concept of “in vivo PHA granule density.” Through the statistical analysis of hundreds of single cells accumulating PHA granules, the distinctive differences of density and localization of PHA granules in the two micro-organisms were found. Furthermore, the team identified the key protein that plays a major role in making the difference that enabled the characteristics of PHA granules in the recombinant E. coli to become similar to those of C. necator.
The research team also presented 3D time-lapse movies showing the actual processes of PHA granule formation combined with cell growth and division. Movies showing the living cells synthesizing and accumulating PHA granules in their native state had never been reported before.
Professor Lee said, “This study provides insights into the morphological and physical characteristics of in vivo PHA as well as the unique mechanisms of PHA granule formation that undergo the phase transition from soluble monomers into the insoluble polymer, followed by granule formation. Through this study, a deeper understanding of PHA granule formation within the bacterial cells is now possible, which has great significance in that a convergence study of biology and physics was achieved. This study will help develop various bioplastics production processes in the future.”
This work was supported by the Technology Development Program to Solve Climate Changes on Systems Metabolic Engineering for Biorefineries (Grants NRF-2012M1A2A2026556 and NRF-2012M1A2A2026557) and the Bio & Medical Technology Development Program (Grant No. 2021M3A9I4022740) from the Ministry of Science and ICT (MSIT) through the National Research Foundation (NRF) of Korea to S.Y.L. This work was also supported by the KAIST Cross-Generation Collaborative Laboratory project.
-PublicationSo Young Choi, Jeonghun Oh, JaeHwang Jung, YongKeun Park, and Sang Yup Lee. Three-dimensional label-free visualization and quantification of polyhydroxyalkanoates in individualbacterial cell in its native state. PNAS(https://doi.org./10.1073/pnas.2103956118)
-ProfileDistinguished Professor Sang Yup LeeMetabolic Engineering and Synthetic Biologyhttp://mbel.kaist.ac.kr/
Department of Chemical and Biomolecular Engineering KAIST
Endowed Chair Professor YongKeun ParkBiomedical Optics Laboratoryhttps://bmokaist.wordpress.com/
Department of PhysicsKAIST
2021.07.28 View 10315 -
Repurposed Drugs Present New Strategy for Treating COVID-19
Virtual screening of 6,218 drugs and cell-based assays identifies best therapeutic medication candidates
A joint research group from KAIST and Institut Pasteur Korea has identified repurposed drugs for COVID-19 treatment through virtual screening and cell-based assays. The research team suggested the strategy for virtual screening with greatly reduced false positives by incorporating pre-docking filtering based on shape similarity and post-docking filtering based on interaction similarity. This strategy will help develop therapeutic medications for COVID-19 and other antiviral diseases more rapidly. This study was reported at the Proceedings of the National Academy of Sciences of the United States of America (PNAS).
Researchers screened 6,218 drugs from a collection of FDA-approved drugs or those under clinical trial and identified 38 potential repurposed drugs for COVID-19 with this strategy. Among them, seven compounds inhibited SARS-CoV-2 replication in Vero cells. Three of these drugs, emodin, omipalisib, and tipifarnib, showed anti-SARS-CoV-2 activity in human lung cells, Calu-3.
Drug repurposing is a practical strategy for developing antiviral drugs in a short period of time, especially during a global pandemic. In many instances, drug repurposing starts with the virtual screening of approved drugs. However, the actual hit rate of virtual screening is low and most of the predicted drug candidates are false positives.
The research group developed effective filtering algorithms before and after the docking simulations to improve the hit rates. In the pre-docking filtering process, compounds with similar shapes to the known active compounds for each target protein were selected and used for docking simulations. In the post-docking filtering process, the chemicals identified through their docking simulations were evaluated considering the docking energy and the similarity of the protein-ligand interactions with the known active compounds.
The experimental results showed that the virtual screening strategy reached a high hit rate of 18.4%, leading to the identification of seven potential drugs out of the 38 drugs initially selected.
“We plan to conduct further preclinical trials for optimizing drug concentrations as one of the three candidates didn’t resolve the toxicity issues in preclinical trials,” said Woo Dae Jang, one of the researchers from KAIST.
“The most important part of this research is that we developed a platform technology that can rapidly identify novel compounds for COVID-19 treatment. If we use this technology, we will be able to quickly respond to new infectious diseases as well as variants of the coronavirus,” said Distinguished Professor Sang Yup Lee.
This work was supported by the KAIST Mobile Clinic Module Project funded by the Ministry of Science and ICT (MSIT) and the National Research Foundation of Korea (NRF). The National Culture Collection for Pathogens in Korea provided the SARS-CoV-2 (NCCP43326).
-PublicationWoo Dae Jang, Sangeun Jeon, Seungtaek Kim, and Sang Yup Lee. Drugs repurposed for COVID-19 by virtual screening of 6,218 drugs and cell-based assay. Proc. Natl. Acad. Sci. U.S.A. (https://doi/org/10.1073/pnas.2024302118)
-ProfileDistinguished Professor Sang Yup LeeMetabolic &Biomolecular Engineering National Research Laboratoryhttp://mbel.kaist.ac.kr
Department of Chemical and Biomolecular EngineeringKAIST
2021.07.08 View 10709 -
Natural Rainbow Colorants Microbially Produced
Integrated strategies of systems metabolic engineering and membrane engineering led to the production of natural rainbow colorants comprising seven natural colorants from bacteria for the first time
A research group at KAIST has engineered bacterial strains capable of producing three carotenoids and four violacein derivatives, completing the seven colors in the rainbow spectrum. The research team integrated systems metabolic engineering and membrane engineering strategies for the production of seven natural rainbow colorants in engineered Escherichia coli strains. The strategies will be also useful for the efficient production of other industrially important natural products used in the food, pharmaceutical, and cosmetic industries.
Colorants are widely used in our lives and are directly related to human health when we eat food additives and wear cosmetics. However, most of these colorants are made from petroleum, causing unexpected side effects and health problems. Furthermore, they raise environmental concerns such as water pollution from dyeing fabric in the textiles industry. For these reasons, the demand for the production of natural colorants using microorganisms has increased, but could not be readily realized due to the high cost and low yield of the bioprocesses.
These challenges inspired the metabolic engineers at KAIST including researchers Dr. Dongsoo Yang and Dr. Seon Young Park, and Distinguished Professor Sang Yup Lee from the Department of Chemical and Biomolecular Engineering. The team reported the study entitled “Production of rainbow colorants by metabolically engineered Escherichia coli” in Advanced Science online on May 5. It was selected as the journal cover of the July 7 issue.
This research reports for the first time the production of rainbow colorants comprising three carotenoids and four violacein derivatives from glucose or glycerol via systems metabolic engineering and membrane engineering. The research group focused on the production of hydrophobic natural colorants useful for lipophilic food and dyeing garments. First, using systems metabolic engineering, which is an integrated technology to engineer the metabolism of a microorganism, three carotenoids comprising astaxanthin (red), -carotene (orange), and zeaxanthin (yellow), and four violacein derivatives comprising proviolacein (green), prodeoxyviolacein (blue), violacein (navy), and deoxyviolacein (purple) could be produced. Thus, the production of natural colorants covering the complete rainbow spectrum was achieved.
When hydrophobic colorants are produced from microorganisms, the colorants are accumulated inside the cell. As the accumulation capacity is limited, the hydrophobic colorants could not be produced with concentrations higher than the limit. In this regard, the researchers engineered the cell morphology and generated inner-membrane vesicles (spherical membranous structures) to increase the intracellular capacity for accumulating the natural colorants. To further promote production, the researchers generated outer-membrane vesicles to secrete the natural colorants, thus succeeding in efficiently producing all of seven rainbow colorants. It was even more impressive that the production of natural green and navy colorants was achieved for the first time.
“The production of the seven natural rainbow colorants that can replace the current petroleum-based synthetic colorants was achieved for the first time,” said Dr. Dongsoo Yang. He explained that another important point of the research is that integrated metabolic engineering strategies developed from this study can be generally applicable for the efficient production of other natural products useful as pharmaceuticals or nutraceuticals. “As maintaining good health in an aging society is becoming increasingly important, we expect that the technology and strategies developed here will play pivotal roles in producing other valuable natural products of medical or nutritional importance,” explained Distinguished Professor Lee.
This work was supported by the "Cooperative Research Program for Agriculture Science & Technology Development (Project No. PJ01550602)" Rural Development Administration, Republic of Korea.
-Publication:Dongsoo Yang, Seon Young Park, and Sang Yup Lee. Production of rainbow colorants by metabolically engineered Escherichia coli. Advanced Science, 2100743.
-Profile Distinguished Professor Sang Yup LeeMetabolic &Biomolecular Engineering National Research Laboratoryhttp://mbel.kaist.ac.kr
Department of Chemical and Biomolecular EngineeringKAIST
2021.06.09 View 8796 -
Identification of How Chemotherapy Drug Works Could Deliver Personalized Cancer Treatment
The chemotherapy drug decitabine is commonly used to treat patients with blood cancers, but its response rate is somewhat low. Researchers have now identified why this is the case, opening the door to more personalized cancer therapies for those with these types of cancers, and perhaps further afield.
Researchers have identified the genetic and molecular mechanisms within cells that make the chemotherapy drug decitabine—used to treat patients with myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) —work for some patients but not others. The findings should assist clinicians in developing more patient-specific treatment strategies.
The findings were published in the Proceedings of the National Academies of Science on March 30.
The chemotherapy drug decitabine, also known by its brand name Dacogen, works by modifying our DNA that in turn switches on genes that stop the cancer cells from growing and replicating. However, decitabine’s response rate is somewhat low (showing improvement in just 30-35% of patients), which leaves something of a mystery as to why it works well for some patients but not for others. To find out why this happens, researchers from the KAIST investigated the molecular mediators that are involved with regulating the effects of the drug.
Decitabine works to activate the production of endogenous retroviruses (ERVs), which in turn induces an immune response. ERVs are viruses that long ago inserted dormant copies of themselves into the human genome. Decitabine in essence, ‘reactivates’ these viral elements and produces double-stranded RNAs (dsRNAs) that the immune system views as a foreign body.
“However, the mechanisms involved in this process, in particular how production and transport of these ERV dsRNAs were regulated within the cell were understudied,” said corresponding author Yoosik Kim, professor in the Department of Chemical and Biomolecular Engineering at KAIST.
“So to explain why decitabine works in some patients but not others, we investigated what these molecular mechanisms were,” added Kim.
To do so, the researchers used image-based RNA interference (RNAi) screening. This is a relatively new technique in which specific sequences within a genome are knocked out of action or “downregulated.” Large-scale screening, which can be performed in cultured cells or within live organisms, works to investigate the function of different genes. The KAIST researchers collaborated with the Institut Pasteur Korea to analyze the effect of downregulating genes that recognize ERV dsRNAs and could be involved in the cellular response to decitabine.
From these initial screening results, they performed an even more detailed downregulation screening analysis. Through the screening, they were able to identify two particular gene sequences involved in the production of an RNA-binding protein called Staufen1 and the production of a strand of RNA that does not in turn produce any proteins called TINCR that play a key regulatory role in response to the drug. Staufen1 binds directly to dsRNAs and stabilizes them in concert with the TINCR.
If a patient is not producing sufficient Staufen1 and TINCR, then the dsRNA viral mimics quickly degrade before the immune system can spot them. And, crucially for cancer therapy, this means that patients with lower expression (activation) of these sequences will show inferior response to decitabine. Indeed, the researchers confirmed that MDS/AML patients with low Staufen1 and TINCR expression did not benefit from decitabine therapy.
“We can now isolate patients who will not benefit from the therapy and direct them to a different type of therapy,” said first author Yongsuk Ku. “This serves as an important step toward developing a patient-specific treatment cancer strategy.”
As the researchers used patient samples taken from bone marrow, the next step will be to try to develop a testing method that can identify the problem from just blood samples, which are much easier to acquire from patients.
The team plans to investigate if the analysis can be extended to patients with solid tumors in addition to those with blood cancers.
-Profile
Professor Yoosik Kim
https://qcbio.kaist.ac.kr/
Department of Chemical and Biomolecular Engineering
KAIST
-Publication
Noncanonical immune response to the inhibition of DNA methylation by Staufen1 via stabilization of endogenous retrovirus RNAs, PNAS
2021.05.24 View 8327 -
Prof. Sang Yup Lee Elected as a Foreign Member of the Royal Society
Vice President for Research Distinguished Professor Sang Yup Lee was elected as a foreign member of the Royal Society in the UK. On May 6, the Society announced the list of distinguished new 52 fellows and 10 foreign members who achieved exceptional contributions to science. Professor Lee and Professor V. Narry Kim from Seoul National University are the first foreign members ever elected from Korea.
The Royal Society, established in 1660, is one of the most prestigious national science academies and a fellowship of 1,600 of the world’s most eminent scientists. From Newton to Darwin, Einstein, Hawking, and beyond, pioneers and paragons in their fields are elected by their peers. To date, there are 280 Nobel prize winners among the fellows.
Distinguished Professor Lee from the Department of Chemical and Biomolecular Engineering at KAIST is one of the Highly Cited Researchers (HCRs) who pioneered systems metabolic engineering and developed various micro-organisms for producing a wide range of fuels, chemicals, materials, and natural compounds.
His seminal scholarship and research career have already been recognized worldwide. He is the first Korean ever elected into the National Academy of Inventors (NAI) in the US and one of 13 scholars elected as an International Member of both the National Academy of Sciences (NAS) and the National Academy of Engineering (NAE) in the US. With this fellowship, he added one more accolade of being the first non-US and British Commonwealth scientist elected into the three most prestigious science academies: the NAS, the NAE, and the Royal Society.
2021.05.07 View 9001 -
Distinguished Professor Sang Yup Lee Honored with Charles D. Scott Award
Vice President for Research Sang Yup Lee received the 2021 Charles D. Scott Award from the Society for Industrial Microbiology and Biotechnology. Distinguished Professor Lee from the Department of Chemical and Biomolecular Engineering at KAIST is the first Asian awardee.
The Charles D. Scott Award, initiated in 1995, recognizes individuals who have made significant contributions to enable and further the use of biotechnology to produce fuels and chemicals. The award is named in honor of Dr. Charles D. Scott, who founded the Symposium on Biomaterials, Fuels, and Chemicals and chaired the conference for its first ten years.
Professor Lee has pioneered systems metabolic engineering and developed various micro-organisms capable of producing a wide range of fuels, chemicals, materials, and natural compounds, many of them for the first time. Some of the breakthroughs include the microbial production of gasoline, diacids, diamines, PLA and PLGA polymers, and several natural products.
More recently, his team has developed a microbial strain capable of the mass production of succinic acid, a monomer for manufacturing polyester, with the highest production efficiency to date, as well as a Corynebacterium glutamicum strain capable of producing high-level glutaric acid. They also engineered for the first time a bacterium capable of producing carminic acid, a natural red colorant that is widely used for food and cosmetics.
Professor Lee is one of the Highly Cited Researchers (HCR), ranked in the top 1% by citations in their field by Clarivate Analytics for four consecutive years from 2017. He is the first Korean fellow ever elected into the National Academy of Inventors in the US and one of 13 scholars elected as an International Member of both the National Academy of Sciences and the National Academy of Engineering in the USA.
The awards ceremony will take place during the Symposium on Biomaterials, Fuels, and Chemicals held online from April 26.
2021.04.27 View 7875 -
Microbial Production of a Natural Red Colorant Carminic Acid
Metabolic engineering and computer-simulated enzyme engineering led to the production of carminic acid, a natural red colorant, from bacteria for the first time
A research group at KAIST has engineered a bacterium capable of producing a natural red colorant, carminic acid, which is widely used for food and cosmetics. The research team reported the complete biosynthesis of carminic acid from glucose in engineered Escherichia coli. The strategies will be useful for the design and construction of biosynthetic pathways involving unknown enzymes and consequently the production of diverse industrially important natural products for the food, pharmaceutical, and cosmetic industries.
Carminic acid is a natural red colorant widely being used for products such as strawberry milk and lipstick. However, carminic acid has been produced by farming cochineals, a scale insect which only grows in the region around Peru and Canary Islands, followed by complicated multi-step purification processes. Moreover, carminic acid often contains protein contaminants that cause allergies so many people are unwilling to consume products made of insect-driven colorants. On that account, manufacturers around the world are using alternative red colorants despite the fact that carminic acid is one of the most stable natural red colorants.
These challenges inspired the metabolic engineering research group at KAIST to address this issue. Its members include postdoctoral researchers Dongsoo Yang and Woo Dae Jang, and Distinguished Professor Sang Yup Lee of the Department of Chemical and Biomolecular Engineering. This study entitled “Production of carminic acid by metabolically engineered Escherichia coli” was published online in the Journal of the American Chemical Society (JACS) on April 2.
This research reports for the first time the development of a bacterial strain capable of producing carminic acid from glucose via metabolic engineering and computer simulation-assisted enzyme engineering. The research group optimized the type II polyketide synthase machinery to efficiently produce the precursor of carminic acid, flavokermesic acid.
Since the enzymes responsible for the remaining two reactions were neither discovered nor functional, biochemical reaction analysis was performed to identify enzymes that can convert flavokermesic acid into carminic acid. Then, homology modeling and docking simulations were performed to enhance the activities of the two identified enzymes. The team could confirm that the final engineered strain could produce carminic acid directly from glucose. The C-glucosyltransferase developed in this study was found to be generally applicable for other natural products as showcased by the successful production of an additional product, aloesin, which is found in aloe leaves.
“The most important part of this research is that unknown enzymes for the production of target natural products were identified and improved by biochemical reaction analyses and computer simulation-assisted enzyme engineering,” says Dr. Dongsoo Yang. He explained the development of a generally applicable C-glucosyltransferase is also useful since C-glucosylation is a relatively unexplored reaction in bacteria including Escherichia coli. Using the C-glucosyltransferase developed in this study, both carminic acid and aloesin were successfully produced from glucose.
“A sustainable and insect-free method of producing carminic acid was achieved for the first time in this study. Unknown or inefficient enzymes have always been a major problem in natural product biosynthesis, and here we suggest one effective solution for solving this problem. As maintaining good health in the aging society is becoming increasingly important, we expect that the technology and strategies developed here will play pivotal roles in producing other valuable natural products of medical or nutritional importance,” said Distinguished Professor Sang Yup Lee.
This work was supported by the Technology Development Program to Solve Climate Changes on Systems Metabolic Engineering for Biorefineries of the Ministry of Science and ICT (MSIT) through the National Research Foundation (NRF) of Korea and the KAIST Cross-Generation Collaborative Lab project; Sang Yup Lee and Dongsoo Yang were also supported by Novo Nordisk Foundation in Denmark.
Publication:
Dongsoo Yang, Woo Dae Jang, and Sang Yup Lee. Production of carminic acid by metabolically engineered Escherichia coli. at the Journal of the American Chemical Society. https://doi.org.10.1021/jacs.0c12406
Profile:
Sang Yup Lee, PhD
Distinguished Professor
leesy@kaist.ac.kr
http://mbel.kaist.ac.kr
Metabolic &Biomolecular Engineering National Research Laboratory
Department of Chemical and Biomolecular Engineering
KAIST
2021.04.06 View 10307 -
Expanding the Biosynthetic Pathway via Retrobiosynthesis
- Researchers reports a new strategy for the microbial production of multiple short-chain primary amines via retrobiosynthesis. -
KAIST metabolic engineers presented the bio-based production of multiple short-chain primary amines that have a wide range of applications in chemical industries for the first time. The research team led by Distinguished Professor Sang Yup Lee from the Department of Chemical and Biomolecular Engineering designed the novel biosynthetic pathways for short-chain primary amines by combining retrobiosynthesis and a precursor selection step.
The research team verified the newly designed pathways by confirming the in vivo production of 10 short-chain primary amines by supplying the precursors. Furthermore, the platform Escherichia coli strains were metabolically engineered to produce three proof-of-concept short-chain primary amines from glucose, demonstrating the possibility of the bio-based production of diverse short-chain primary amines from renewable resources. The research team said this study expands the strategy of systematically designing biosynthetic pathways for the production of a group of related chemicals as demonstrated by multiple short-chain primary amines as examples.
Currently, most of the industrial chemicals used in our daily lives are produced with petroleum-based products. However, there are several serious issues with the petroleum industry such as the depletion of fossil fuel reserves and environmental problems including global warming. To solve these problems, the sustainable production of industrial chemicals and materials is being explored with microorganisms as cell factories and renewable non-food biomass as raw materials for alternative to petroleum-based products. The engineering of these microorganisms has increasingly become more efficient and effective with the help of systems metabolic engineering – a practice of engineering the metabolism of a living organism toward the production of a desired metabolite. In this regard, the number of chemicals produced using biomass as a raw material has substantially increased.
Although the scope of chemicals that are producible using microorganisms continues to expand through advances in systems metabolic engineering, the biological production of short-chain primary amines has not yet been reported despite their industrial importance. Short-chain primary amines are the chemicals that have an alkyl or aryl group in the place of a hydrogen atom in ammonia with carbon chain lengths ranging from C1 to C7. Short-chain primary amines have a wide range of applications in chemical industries, for example, as a precursor for pharmaceuticals (e.g., antidiabetic and antihypertensive drugs), agrochemicals (e.g., herbicides, fungicides and insecticides), solvents, and vulcanization accelerators for rubber and plasticizers. The market size of short-chain primary amines was estimated to be more than 4 billion US dollars in 2014.
The main reason why the bio-based production of short-chain primary amines was not yet possible was due to their unknown biosynthetic pathways. Therefore, the team designed synthetic biosynthetic pathways for short-chain primary amines by combining retrobiosynthesis and a precursor selection step. The retrobiosynthesis allowed the systematic design of a biosynthetic pathway for short-chain primary amines by using a set of biochemical reaction rules that describe chemical transformation patterns between a substrate and product molecules at an atomic level.
These multiple precursors predicted for the possible biosynthesis of each short-chain primary amine were sequentially narrowed down by using the precursor selection step for efficient metabolic engineering experiments.
“Our research demonstrates the possibility of the renewable production of short-chain primary amines for the first time. We are planning to increase production efficiencies of short-chain primary amines. We believe that our study will play an important role in the development of sustainable and eco-friendly bio-based industries and the reorganization of the chemical industry, which is mandatory for solving the environmental problems threating the survival of mankind,” said Professor Lee.
This paper titled “Microbial production of multiple short-chain primary amines via retrobiosynthesis” was published in Nature Communications. This work was supported by the Technology Development Program to Solve Climate Changes on Systems Metabolic Engineering for Biorefineries from the Ministry of Science and ICT through the National Research Foundation (NRF) of Korea.
-Publication
Dong In Kim, Tong Un Chae, Hyun Uk Kim, Woo Dae Jang, and Sang Yup Lee. Microbial production of multiple short-chain primary amines via retrobiosynthesis. Nature Communications ( https://www.nature.com/articles/s41467-020-20423-6)
-Profile
Distinguished Professor Sang Yup Lee
leesy@kaist.ac.kr
Metabolic &Biomolecular Engineering National Research Laboratory
http://mbel.kaist.ac.kr
Department of Chemical and Biomolecular Engineering
KAIST
2021.01.14 View 10240 -
Researchers Report Longest-lived Aqueous Flow Batteries
New technology to overcome the life limit of next-generation water-cell batteries
A research team led by Professor Hee-Tak Kim from the Department of Chemical and Biomolecular Engineering has developed water-based zinc/bromine redox flow batteries (ZBBs) with the best life expectancy among all the redox flow batteries reported by identifying and solving the deterioration issue with zinc electrodes.
Professor Kim, head of the Advanced Battery Center at KAIST's Nano-fusion Research Institute, said, "We presented a new technology to overcome the life limit of next-generation water-cell batteries. Not only is it cheaper than conventional lithium-ion batteries, but it can contribute to the expansion of renewable energy and the safe supply of energy storage systems that can run with more than 80 percent energy efficiency."
ZBBs were found to have stable life spans of more than 5,000 cycles, even at a high current density of 100 mA/cm2. It was also confirmed that it represented the highest output and life expectancy compared to Redox flow batteries (RFBs) reported worldwide, which use other redox couples such as zinc-bromine, zinc-iodine, zinc-iron, and vanadium.
Recently, more attention has been focused on energy storage system (ESS) that can improve energy utilization efficiency by storing new and late-night power in large quantities and supplying it to the grid if necessary to supplement the intermittent nature of renewable energy and meet peak power demand.
However, lithium-ion batteries (LIBs), which are currently the core technology of ESSs, have been criticized for not being suitable for ESSs, which store large amounts of electricity due to their inherent risk of ignition and fire. In fact, a total of 33 cases of ESSs using LIBs in Korea had fire accidents, and 35% of all ESS facilities were shut down. This is estimated to have resulted in more than 700 billion won in losses.
As a result, water-based RFBs have drawn great attention. In particular, ZBBs that use ultra-low-cost bromide (ZnBr2) as an active material have been developed for ESSs since the 1970s, with their advantages of high cell voltage, high energy density, and low price compared to other RFBs. Until now, however, the commercialization of ZBBs has been delayed due to the short life span caused by the zinc electrodes. In particular, the uneven "dendrite" growth behavior of zinc metals during the charging and discharging process leads to internal short circuits in the battery which shorten its life.
The research team noted that self-aggregation occurs through the surface diffusion of zinc nuclei on the carbon electrode surface with low surface energy, and determined that self-aggregation was the main cause of zinc dendrite formation through quantum mechanics-based computer simulations and transmission electron microscopy. Furthermore, it was found that the surface diffusion of the zinc nuclei was inhibited in certain carbon fault structures so that dendrites were not produced.
Single vacancy defect, where one carbon atom is removed, exchanges zinc nuclei and electrons, and is strongly coupled, thus inhibiting surface diffusion and enabling uniform nuclear production/growth. The research team applied carbon electrodes with high density fault structure to ZBBs, achieving life characteristics of more than 5,000 cycles at a high charge current density (100 mA/cm2), which is 30 times that of LIBs.
This ESS technology, which can supply eco-friendly electric energy such as renewable energy to the private sector through technology that can drive safe and cheap redox flow batteries for long life, is expected to draw attention once again.
Publication:
Ju-Hyuk Lee, Riyul Kim, Soohyun Kim, Jiyun Heo, Hyeokjin Kwon, Jung Hoon Yang, and Hee-Tak Kim. 2020. Dendrite-free Zn electrodeposition triggered by interatomic orbital hybridization of Zn and single vacancy carbon defects for aqueous Zn-based flow batteries. Energy and Environmental Science, 2020, 13, 2839-2848.
Link to download the full-text paper:http://xlink.rsc.org/?DOI=D0EE00723D
Profile: Prof. Hee-Tak Kimheetak.kim@kaist.ac.krhttp://eed.kaist.ac.krAssociate ProfessorDepartment of Chemical & Biomolecular EngineeringKAIST
2020.12.16 View 11028 -
A Comprehensive Review of Biosynthesis of Inorganic Nanomaterials Using Microorganisms and Bacteriophages
There are diverse methods for producing numerous inorganic nanomaterials involving many experimental variables. Among the numerous possible matches, finding the best pair for synthesizing in an environmentally friendly way has been a longstanding challenge for researchers and industries.
A KAIST bioprocess engineering research team led by Distinguished Professor Sang Yup Lee conducted a summary of 146 biosynthesized single and multi-element inorganic nanomaterials covering 55 elements in the periodic table synthesized using wild-type and genetically engineered microorganisms. Their research highlights the diverse applications of biogenic nanomaterials and gives strategies for improving the biosynthesis of nanomaterials in terms of their producibility, crystallinity, size, and shape.
The research team described a 10-step flow chart for developing the biosynthesis of inorganic nanomaterials using microorganisms and bacteriophages. The research was published at Nature Review Chemistry as a cover and hero paper on December 3.
“We suggest general strategies for microbial nanomaterial biosynthesis via a step-by-step flow chart and give our perspectives on the future of nanomaterial biosynthesis and applications. This flow chart will serve as a general guide for those wishing to prepare biosynthetic inorganic nanomaterials using microbial cells,” explained Dr.Yoojin Choi, a co-author of this research.
Most inorganic nanomaterials are produced using physical and chemical methods and biological synthesis has been gaining more and more attention. However, conventional synthesis processes have drawbacks in terms of high energy consumption and non-environmentally friendly processes. Meanwhile, microorganisms such as microalgae, yeasts, fungi, bacteria, and even viruses can be utilized as biofactories to produce single and multi-element inorganic nanomaterials under mild conditions.
After conducting a massive survey, the research team summed up that the development of genetically engineered microorganisms with increased inorganic-ion-binding affinity, inorganic-ion-reduction ability, and nanomaterial biosynthetic efficiency has enabled the synthesis of many inorganic nanomaterials.
Among the strategies, the team introduced their analysis of a Pourbaix diagram for controlling the size and morphology of a product. The research team said this Pourbaix diagram analysis can be widely employed for biosynthesizing new nanomaterials with industrial applications.Professor Sang Yup Lee added, “This research provides extensive information and perspectives on the biosynthesis of diverse inorganic nanomaterials using microorganisms and bacteriophages and their applications. We expect that biosynthetic inorganic nanomaterials will find more diverse and innovative applications across diverse fields of science and technology.”
Dr. Choi started this research in 2018 and her interview about completing this extensive research was featured in an article at Nature Career article on December 4.
-ProfileDistinguished Professor Sang Yup Lee leesy@kaist.ac.krMetabolic &Biomolecular Engineering National Research Laboratoryhttp://mbel.kaist.ac.krDepartment of Chemical and Biomolecular EngineeringKAIST
2020.12.07 View 8897 -
Engineered C. glutamicum Strain Capable of Producing High-Level Glutaric Acid from Glucose
An engineered C. glutamicum strain that can produce the world’s highest titer of glutaric acid was developed by employing systems metabolic engineering strategies
A metabolic engineering research group at KAIST has developed an engineered Corynebacterium glutamicum strain capable of producing high-level glutaric acid without byproducts from glucose. This new strategy will be useful for developing engineered micro-organisms for the bio-based production of value-added chemicals.
Glutaric acid, also known as pentanedioic acid, is a carboxylic acid that is widely used for various applications including the production of polyesters, polyamides, polyurethanes, glutaric anhydride, 1,5-pentanediol, and 5-hydroxyvaleric acid. Glutaric acid has been produced using various petroleum-based chemical methods, relying on non-renewable and toxic starting materials. Thus, various approaches have been taken to biologically produce glutaric acid from renewable resources. Previously, the development of the first glutaric acid producing Escherichia coli by introducing Pseudomonas putida genes was reported by a research group from KAIST, but the titer was low. Glutaric acid production by metabolically engineered Corynebacterium glutamicum has also been reported in several studies, but further improvements in glutaric acid production seemed possible since C. glutamicum has the capability of producing more than 130 g/L of L-lysine.
A research group comprised of Taehee Han, Gi Bae Kim, and Distinguished Professor Sang Yup Lee of the Department of Chemical and Biomolecular Engineering addressed this issue. Their research paper “Glutaric acid production by systems metabolic engineering of an L-lysine-overproducing Corynebacterium glutamicum” was published online in PNAS on November 16, 2020.
This research reports the development of a metabolically engineered C. glutamicum strain capable of efficiently producing glutaric acid, starting from an L-lysine overproducer. The following novel strategies and approaches to achieve high-level glutaric acid production were employed. First, metabolic pathways in C. glutamicum were reconstituted for glutaric acid production by introducing P. putida genes. Then, multi-omics analyses including genome, transcriptome, and fluxome were conducted to understand the phenotype of the L-lysine overproducer strain. In addition to systematic understanding of the host strain, gene manipulation targets were predicted by omics analyses and applied for engineering C. glutamicum, which resulted in the development of an engineered strain capable of efficiently producing glutaric acid. Furthermore, the new glutaric acid exporter was discovered for the first time, which was used to further increase glutaric acid production through enhancing product excretion. Last but not least, culture conditions were optimized for high-level glutaric acid production. As a result, the final engineered strain was able to produce 105.3 g/L glutaric acid, the highest titer ever reported, in 69 hours by fed-batch fermentation.
Professor Sang Yup Lee said, “It is meaningful that we were able to develop a highly efficient glutaric acid producer capable of producing glutaric acid at the world’s highest titer without any byproducts from renewable carbon sources. This will further accelerate the bio-based production of valuable chemicals in pharmaceutical/medical/chemical industries.” This research was supported by the Bio & Medical Technology Development Program of the National Research Foundation and funded by the Ministry of Science and ICT.
-Profile
Distinguished Professor Sang Yup Lee
leesy@kaist.ac.kr
http://mbel.kaist.ac.kr
Department of Chemical and Biomolecular Engineering
KAIST
2020.11.17 View 6206 -
E. coli Engineered to Grow on CO₂ and Formic Acid as Sole Carbon Sources
- An E. coli strain that can grow to a relatively high cell density solely on CO₂ and formic acid was developed by employing metabolic engineering. -
Most biorefinery processes have relied on the use of biomass as a raw material for the production of chemicals and materials. Even though the use of CO₂ as a carbon source in biorefineries is desirable, it has not been possible to make common microbial strains such as E. coli grow on CO₂.
Now, a metabolic engineering research group at KAIST has developed a strategy to grow an E. coli strain to higher cell density solely on CO₂ and formic acid. Formic acid is a one carbon carboxylic acid, and can be easily produced from CO₂ using a variety of methods. Since it is easier to store and transport than CO₂, formic acid can be considered a good liquid-form alternative of CO₂.
With support from the C1 Gas Refinery R&D Center and the Ministry of Science and ICT, a research team led by Distinguished Professor Sang Yup Lee stepped up their work to develop an engineered E. coli strain capable of growing up to 11-fold higher cell density than those previously reported, using CO₂ and formic acid as sole carbon sources. This work was published in Nature Microbiology on September 28.
Despite the recent reports by several research groups on the development of E. coli strains capable of growing on CO₂ and formic acid, the maximum cell growth remained too low (optical density of around 1) and thus the production of chemicals from CO₂ and formic acid has been far from realized.
The team previously reported the reconstruction of the tetrahydrofolate cycle and reverse glycine cleavage pathway to construct an engineered E. coli strain that can sustain growth on CO₂ and formic acid. To further enhance the growth, the research team introduced the previously designed synthetic CO₂ and formic acid assimilation pathway, and two formate dehydrogenases.
Metabolic fluxes were also fine-tuned, the gluconeogenic flux enhanced, and the levels of cytochrome bo3 and bd-I ubiquinol oxidase for ATP generation were optimized. This engineered E. coli strain was able to grow to a relatively high OD600 of 7~11, showing promise as a platform strain growing solely on CO₂ and formic acid.
Professor Lee said, “We engineered E. coli that can grow to a higher cell density only using CO₂ and formic acid. We think that this is an important step forward, but this is not the end. The engineered strain we developed still needs further engineering so that it can grow faster to a much higher density.”
Professor Lee’s team is continuing to develop such a strain. “In the future, we would be delighted to see the production of chemicals from an engineered E. coli strain using CO₂ and formic acid as sole carbon sources,” he added.
-Profile:Distinguished Professor Sang Yup Leehttp://mbel.kaist.ac.krDepartment of Chemical and Biomolecular EngineeringKAIST
2020.09.29 View 9528